Development of a novel Yeast Surface Display system for engineering of small proteins

Abstract

Yeast surface display has been an emerging technology for protein engineering and characterization in hope to achieve higher protein affinity and stability. Yeast serves as an ideal model for eukaryotic organism as its expression system is comparable with mammalian cells, which offers several advantages over prokaryotic processing, such as post-translational modifications and proper folding within the endoplasmic reticulum. The most common technology is based on the interaction between a receptor on the Saccharomyces cerevisiae membrane, Aga1, with α-agglutinin mating protein, Aga2, via disulfate bond, which allows the display of C-terminal fused proteins of interest such as: antibodies and labelling agents. Currently there are limitations regarding the universality of the YSD system, its application being strictly related to the yeast strain of interest and the number of libraries is between 107 to 109, inferior to other display methodologies. The aim of the thesis is to define a novel YSD where the promotor, secretion signal, and anchor protein combination depend on the cell type and proteins of interest. The strategy used was to identify a common surface anchor for yeast cell belonging to different fungi families, to which a ligand can be chemically attached, thus, to enable the recognition of genetically engineered proteins.